Understanding cardiac progenitor pathways is relevant for both congenital and adult heart disease. The LIM homeodomain transcription factor Islet (Isl1) plays a pivotal role in cardiac progenitors during development, and is also expressed in postnatal cardiac progenitors found in rodent and human heart. During cardiogenesis, Isl1 is required for proliferation, survival, and migration of second heart field progenitors, and is expressed both within cardiogenic mesoderm and pharyngeal endoderm. It is not known whether Isl1 is required within mesoderm or endoderm for early heart development. Although isl1 is downregulated in the great majority of progenitors as they enter the heart, recent studies have demonstrated persistent Isl1 expression in distinct subdomains within the heart, including myocardial lineages of the outflow tract and sinoatrial node. Persistence of Isl1 in selected differentiated myocardial lineages suggests later requirements for Isl1 which have not been explored. The hypothesis to be tested in this proposal is that Isl1 is required in endoderm, cardiogenic mesoderm, and in subdomains of differentiated cardiomyocytes for specific aspects of heart development and function, and that defining direct or indirect targets of Isl1 will give mechanistic insights into Isl1 action during cardiogenesis. The Specific Aims of this proposal are: (1) To determine tissue specific requirements for Isl1 in mesodermal progenitors or pharyngeal endoderm, utilizing Mesp1-cre or Foxa2- inducible cre, and a floxed allele of isl1;(2) To investigate a potential role for Isl1 in differentiated myocardial lineages by ablation of isl1 in differentiated myocardial lineages utilizing an anterior heart field specific-cre (MEF2c-cre) or a conduction system specific-inducible cre (HCN4-CreERT2), and a floxed allele of isl1;and (3) To gain insight into mechanisms by which Isl1 controls cell behaviors, by identifying direct and indirect downstream targets of Isl1 during early cardiogenesis. Results of these studies will give mechanistic insight into pathways by which Isl1 controls cell behaviors and morphogenesis during heart development. PUBLIC HEALTH RELEVANCE Isl1 is a transcription factor which marks undifferentiated cardiac progenitors and is required for their proliferation, differentiation and migration. Isl1 is also expressed in pharyngeal endoderm, which is adjacent to cardiogenic mesoderm and acts as an inducer. The proposed studies will provide mechanistic insight as to where, when, and how Isl1 is required to effect cell behaviors during cardiogenesis.